BIOCHEMISTRY GENERAL MEDICINE HORMONAL REGULATION

Transcription

1 BIOCHEMISTRY GENERAL MEDICINE HORMONAL REGULATION RNDr. Zdeněk DVOŘÁK, PhD. Department of Medical Chemistry and Biochemistry Faculty of Medicine, Palacky University Olomouc

2 HORMONAL REGULATION Compounds involved in the co-ordination of metabolic activities of various organs and tissues cellular signalling; signal transduction HORMONES NEUROTRANSMITTERS GROWTH FACTORS CYTOKINES Extracellular signalling substances Synthesis in one class of cell transmission TARGET CELLS HORMONES Synthetized by specific tissues ENDOCRINE GLANDS Secreted directly into the BLOODSTREAM and carried to their sites of action Specifically alter METABOLIC ACTIVITIES of TARGET CELLS (remote from secretory organ) Active at very low concentrations (pm µm) Rapidly metabolized SHORT-LIVED EFFECT

3 SIGNAL TRANSDUCTION SIGNAL HORMONE RECEPTOR TRANSDUCTOR EFFECTOR ULTIMATE CELLULAR RESPONSE SECOND MESSENGER

4 INTEGRATION AND CONTROL OF METABOLIC PROCESSES Senzory inputs from the environment CENTRAL NERVOUS SYSTEM Primary target Hypothalamus Anterior pituitary Posterior pituitary Thyrotropin Corticotropin Secondary target Somatotropin Luteinizing hormone Folicule Stimulating hormone Prolactin Vasopresin Oxytocin Thyroid Adrenal cortex Adrenal medula Pancreatic Islet cells Ovary Testis Thyroxine Triiodothyronne Cortisol Corticosterone Aldosterone Epinephrine Norepinephrine Insulin Glucagon Somatostatin Progesterone Estradiol Testosterone Muscles Liver Many tissues Liver Muscles Heart Liver Muscles Reproductive organs Mammary glands Smooth Muscle; Mammary glands Arterioles Ultimate target

5 CLASSIFICATION OF HORMONES ACCORDING TO STRUCTURE 1. Derived from aminoacids - epinephrine (adrenaline) - nor-epinephrine - thyroxine - triiodthyronine epinephrine 2. Peptides and aminoacids - insulin; glucagon - liberins; oxytocin; vasopresin - ADH; ACTH oxytocin 3. Steroids - cortisol; aldosterone - progesterone; estradiol; testosterone 4. Eicosanoids - prostaglandins; leucotrienes - prostacyclines; thromboxanes estradiol arachidonic acid

6 CLASSIFICATION OF HORMONES ACCORDING TO MODE OF ACTION 1. Hormone DOES NOT TRAVERSE plasma membrane of a target cell amino acids; peptides hydrophylic compounds binding of hormone to the RECEPTOR at the CELL SURFACE hormonal response inside the cell changes in AFFINITY of key proteins/enzymes ACTION THROUGH: - second messengers - activation of enzyme activity of cytosolic receptor domain - opening ion channels SHORT-TERM action; ULTRA-RAPID response 2. Hormone TRAVERSES plasma membrane of a target cell steroid and thyroid hormones lipophylic compounds binding of hormone to the INTRACELLULAR RECEPTOR HORMONE-RECEPTOR complex binds to DNA and triggers TRASCRIPTION of specific genes changes in LEVEL of key proteins/enzymes LONGTERM action; SLOW response

7 RECEPTORS proteins located in plasma membrane or in interior of the cell 2 binding sites - hormone binding site - a component of signal transduction system Peptides, AA HORMONE BINDING (saturable; M.-M. kinetics) P GR GR P GRE TR RAR Extensive conformation change of receptor thyroid GR GR retinoids ACTIVATION OF SECOND BINDING SITE steroids

8 HORMONE Several classes of receptor Identical hormone binding site ADRENERGIC RECEPTORS α1 coupled to phosphatidyl inositol cascade α2, β1, β2 coupled to adenylate cyclase cascade α1 salivary gland K + ; H 2 O secretion α2 pancreatic β cells secretion - muscle glycogenolysis β1 heart rate; contraction force - adipocyte lipolysis β2 liver glycogenolysis Different second binding site Various physiological effects (different tissue) increase in receptor density in the cell = increased cellular response

9 GTP- binding proteins = G - proteins guanyl-nucleotides proteins TRANSDUCTORS = carriers of excitation signal from RECEPTOR to EFFECTOR within plasma membrane peripheral membrane proteins (located on cytosolic side) TRIMER = it consists of α, β and γ subunits α subunit is binding site for GDP/GDP; GTPase activity large extracellular ligand binding domain formation of hormone-receptor complex conformation change of the receptor transduction of signal inside the cell receptor interacts with G-protein exchange of GDP for GTP in α-subunit

10 release of α-(gtp) subunit activation of effector hydrolysis of GTP to GDP release of α-gdp from effector re-association with β and γ subunits

11 G - protein signalling G - proteins G S - stimulatory G i - inhibitory Interaction with different receptors Inhibition of GTPase activity of α-gtp subunit results in irreversibile inactivation of effector and consequently in uncontrolled cellular response Example: Vibrio cholerae toxin inhibits GTPase activity in enterocytes that in turn leads to permanent activation of the effector (adenylate cyclase). As the result; the secretion of Na+ and H 2 O is uncontrolled and severe dehydratation of organism is developed. Stimulus Receptor G-prot. Effector Response Epinephrine β-adrenergic r. Gs Adenylate cyclase Glycogen breakdown Serotonin Serotonin r. Gs Adenylate cyclase Behavioral sensitization Light Rhodopsin Transducin cgmp phosphodiest. Visual excitation IgE-antigen complex Mast cell IgE r. G PLC Phospholipase C Secretion f-met peptide Chemotactic r. G PLC Phospholipase C Chemotaxis Acetylcholine Muscarinic r. Gk Potassium channel Slowing pacemaker activity

12 SECOND MESSENGERS amplified intracellular signals products of activated effector action small molecules or ions allosteric effectors camp - cyclic adenosine monophosphate cgmp - cyclic guanosine monophosphate DAG - 1,2-diacylglycerol IP3 - inositol-1,4,5-triphosphate Ca2+ - calcium (free or bound to calmodulin) SIGNAL TRANSDUCTION SYSTEMS ADENYLATE CYCLASE system PHOSPHATIDYLINOSITOL system TYROSINE KINASE system GUANYLATE CYCLASE system STEROID/THYROID/RETINOID SIGNALLING

13 ADENYLATE CYCLASE (AC) SYSTEM AC is activated via G-protein effector = membrane AC AC converts ATP to camp elevation of cytosolic camp camp is second messenger camp action camp is allosteric activator of proteinkinases activated proteinkinases phosphorylate target proteins phosphorylated proteins (enzymes) have altered functions cellular response degradation of storage fuels HCl secretion by gastric mucosa aggregation of blood platelets

14 Obrázek IX/5

15 ACTIVATION OF PROTEIN KINASE A (PKA) BY camp PKA - 2 catalytic and 2 regulatory subunits camp binds to regulatory subunits heterotetramer dissociates active catalytic subunits of PKA are released PKA phosphorylates cytosolic proteins PKA may enter nucleus and phosphorylate TFs phosphorylated TFs recruit co-activators gene expression is triggered

16 camp signalling Hormones using camp as second messenger Calcitonin Chorionic gonadotropin Corticotropin Epinephrine Follicle-stimulating hormone Glucagon Luteinizing hormone Nor-epinephrine Lipotropin Melanocyte-stimulating hormone Parathyroid hormone Thyroid-stimulating hormone Vasopressin

17 Extracellular side GUANYLATE CYCLASE SYSTEM Peptide hormone (natriuretic factor) Cytosolic side P P P P hydrolysis by phosphodiesterases GTP P P P P inactivation loss of cellular response cgmp inhibited by VIAGRA prolonged cell response cellular response e.g. NO synthesis P

18 PHOSPHATIDYL INOSITOL PHOSPHATE (PIP) SYSTEM principal molecule in the signalling is phosphatidyl inositol-4,5-bis-phosphate (PIP2) membrane receptor trasmits signal and activate effector PI3 kinase PI3K phosphorylates PIP2 to PIP3 phosphatidyl inositol-3,4,5-tris-phosphate (PIP3)

19 PIP3 recruits tyrosin kinase (BTK) and phospholipase C (PLC) BTK phosphorylates PLC activated PLC cleaves PIP2 to 2 fragments 2 second messengers!!! IP3 = inositol-1,4,5-triphophate DAG = 1,2-diacylglycerol

20 IP3 binds calcium channel in ER channels open and releases Ca 2+ in cytosol Ca 2+ binds protein kinase C (PKC) PKC then goes to plasma membrane PKC is activated when both, i.e. calcium and DAG are bound

21 PKC phosporylates its target proteins PIP Important role in control of cell division and proliferation Protein Kinase C Phosphorylates variety of target proteins; e.g.: insulin receptor; glucose carrier; CYP P450; tyrosine hydroxylase etc. PIP system is activated via variety of stimuli; e.g. ACTH, epinephrine, Neurotransmitters, growth factors, antigens Inactivation of PIP system: IP3 IP2 PIP system mediates variety of effects; e.g. Glycogenolysis in liver cells; Histamine secretion by mast cells; Serotonine release by blood platelets; Insulin secretion by pancreatic islet cells; Smoth muscle contraction; Epinephrine secretion by adrenal chromaffin cells; visual transduction

22 Endoplasmic reticulum Ca 2+ Ca 2+ Ca 2+ Ca2+ is released from ER in response to hormones or neurotransmitters Calmodulin-Ca2+ complex is an essential component of many Ca2+ dependent enzymes Ca 2+ calmodulin Transinet increase of intracellular Ca2+ favors formation of complex Ca 2+ Ca 2+ calmodulin Complex Calmodulin-Ca2+ Ca 2+ Ca 2+ Ca 2+ Ca 2+ calmodulin active enzyme Ca 2+ Ca 2+ inactive enzyme substrate product

23 TYROSINE KINASE (TK) SYSTEM membrane receptor contains effector domain effector = tyrosine kinase (TK); protein kinase specific for phosphorylation of tyrosine residue second messenger = O? x IRS extracellular signals insulin, nerve growth factor; epidermal growth factor insulin ATP ADP autophosphorylation P P catalytic activity of TK switched on by insuline binding enhanced activity independent of insuline binding P

24 TYROSINE KINASE (TK) SYSTEM hormone binds TK = H-R complex = TK activation TK phosphorylates target proteins (e.g. phosphatases; amino acid transporter; glucose transporter etc.) = CELLULAR RESPONSE autophosphorylation of TK ensures that TK activity is switched on even in absence of hormone = long term effect of insulin insulin has general growth-promoting properties it acts as metabolic activator and growth factor almost in all cells in the body it acts SHORT-TERM effects = polysaccharide and fat synthesis LONG-TERM effects = nucleic acud and protein synthesis

25 OUTSIDE CELL Nitric oxide Hormonal signals membrane receptors INSIDE CELL G-prot G-prot G-prot G-prot Tyrosin kinase Cyclic AMP Cyclic GMP Ca2+ DAG Proteins IP3 second messengers PKA PKG Calmodulin PKC Ser/Thr kinases Protein substrates Multifunctional kinases Dedicated kinases Protein substrates Protein substrates Other phospholipases

26 STEROID AND THYROID HORMONES ACTION steroid/thyroid receptors intracellular localization cytosolic - glucocorticoid; estrogen; androgen; progesterone; mineralocorticoid (undergo nucle-cytosolic traslocation) nuclear retinoid (RARs, RXRs), vitamine D, thyroid no second messenger DNA is the effector binding to DNA triggering gene expression = cellular response enhanced synthesis of specific proteins Hormone travers plasma membrane Hormone + Receptor Nuclear translocation HR complex binding to DNA cellular response Outside of cell cytosol or nucleus

27 Free steroid hormone Plasma-bound steroid hormone Biological response hsp NEW PROTEIN HR HR HR Protein synthesis HR HR HR transcription HR mrna mrna translation DNA

28 ligand Transcription Iniciation komplex DNA HR HR Co-repressor Co-activator RNA transcription PROMOTER REGION Hormone response element TR, VDR and RARs regulation of transcription: Hormone receptor (HR) is dimerized and bound to DNA at hormone response element site. Without the ligand, transcription is inactive due to the interaction of HR with co-repressor. When hormone binds to HR, the bound co-repressor dissociates leading to an interaction between co-activator and HR. These regulatory changes result in increased transcription.

29 Target organs and genes for steroid and thyroid hormones Hormone class Target organ Target gene Glucocorticoids Liver tyrosine aminotransferase tryptophan oxygenase α-fetoprotein (down-regulation) metallothionein Liver, Retina glutamine synthetase Kidney Phophoenolpyruvate carboxykinase (PEPCK) Oviduct Ovalbumin Pituitary Pro-opiomelanocortin Estrogens Oviduct Ovalbumin; Lysozyme Progesterone Oviduct Ovalbumin; Ovidin Uterus Uteroglobine Androgens Prostate Aldolase Kidney β-glucuronidase Oviduct Albumin 1,25-dihydroxyvimin D3 Intestine Calcium-binding protein Thyroid hormones Liver Carbamoyl phosphyte synthetase; Malic enzyme Pituitary Growth hormone; Prolactin (down-regulation)

30 HORMONE ACTION AND CARCINOGENESIS Alteration of structure of any component in signal transduction system Loss of metabolic control in a cell Transformation of normal cell G-proteins insulin receptor thyroid/steroid hormone receptors DAG Analogs encoded by specific VIRAL ONCOGENES = proteins which bind hormone (HR complex) but LACK SWITCH OFF mechanism (lack of GTPase activity; permanent TK activity; tight binding of hormone Analogs (PHORBOL ESTERS) = cause activation of protein kinase C = stimulation of tumor formation (mainly in presence of carcinogen) Loss of metabolic control; uncontrolled cell growth; transformation of a normal cell to a cancer cell